A
thermolysin-like metalloendopeptidase, optimally active at a neutral pH, was identified in human serum. The
enzyme cleaves the synthetic substrate glutaryl-Ala-Ala-Phe-2-naphthylamide at the
Ala-Phe bond. Activity was determined by measuring the rate of formation of Phe-2-naphthylamide in a coupled
enzyme assay in the presence of excess
aminopeptidase M.
2-Naphthylamine released during the reaction was determined by a diazotization procedure.
Enzyme activity is not affected by inhibitors of
serine,
thiol, or carboxyl
proteases, but is sensitive to inhibition by
metal chelators such as
EDTA and
o-phenanthroline. Dialysis against
EDTA leads to loss of activity, which can be fully restored by
zinc and
cobalt ions. The serum
enzyme closely resembles a membrane-bound
metalloendopeptidase (EC 3.4.24.11) abundant in lung, spleen, and kidney in that both
enzymes are inhibited by the same active-site-directed inhibitors. In addition, an antiserum obtained against the
metalloendopeptidase from rabbit kidney shows strong cross-reactivity with the serum
enzyme.
Metalloendopeptidase activity was measured in 150 controls and in 95 patients with
sarcoidosis; the two groups had significantly different
enzyme activities (p less than 0.001). The mean
enzyme activity in the
sarcoidosis group was more than threefold higher than that of the control group. The mean
enzyme activity for patients with active disease was more than double that of patients with inactive disease and more than four times that of controls (p less than 0.001). This is noteworthy because
angiotensin converting enzyme, a
zinc-
dipeptidyl carboxypeptidase with a mechanism of action similar to that of the
metalloendopeptidase, has also been reported to be increased in the serum of patients with active
sarcoidosis.
Enzyme activity in patients with active
tuberculosis, primary
pulmonary neoplasms, and idiopathic interstitial
pulmonary fibrosis did not differ significantly from that of controls.